Protein structure modification to elucidate the processes of energy transduction in oxygenation, component binding and interaction during the catalytic cycle: NADH plus CH2 plus O2 to NAD ion plus CHOH plus OH2 is projected. Our efforts to reach the details concerning coupled energy transport and oxygen reduction, have led to the isolation of the only purified mixed function oxygenase system to date. This three component methylene hydroxylase from the bacterium Pseudomonas putida catalyzes the conversion of D (plus) camphor to the 5-exo-alcohol in the first of an eleven step degradation to isobutyrate and consists of three separable, homogeneous proteins: (1) an NADH linked FAD flavoprotein reductase, (2) a two-iron two-sulfur redoxin, and (3) a general oxygen binding b type cytochrome P450 cam. Through an intense collaboration of biochemists, molecular geneticists, physicists and organic chemists, we have made great progress in establishing the physical, chemical and biological properties of the hydroxylase system and elucidating the general reaction scheme. Such a characterization has been valuable to the establishment of the analogous parameters from inhomogeneous preparations of mammalian P450 cytochrome. Two general problems, however, remain: What is the detailed mechanism of enzymatic oxygen reduction in mixed function oxidation, and what is the pathway and method by which energy is transferred from NADH and segregated in the final end products? We believe that these two central and historic problems of biology will yield to investigative efforts designed to selectively alter or modify the individual amino acid side chains and substrates in such a way as to yield uneffected, diminished, or destroyed catalytic activity. In this manner the specific residues responsible for chemical catalysis, electron transport, multienzyme aggregation and substrate binding can be revealed. We wish to probe the detailed structure of the intermediates, the enzyme active sites, and the protein-protein interactions during the cycle. Isotope replacement, affinity labels and fluorescent probes will be applied to specific amino acid residues. This is, in our view, the next logical step in the exploration and ultimate understanding of two prime processes of the storage and transmission of information and energy.